Use of glutathione synthesis stimulating compounds in reducing insulin resistance

ABSTRACT

There is provided a method of reducing insulin resistance in a mammalian patient comprising selecting a patient suffering from insulin resistance and administering a compound which increases hepatic glutathione and a compound which increases hepatic nitric oxide.

This application claims priority of invention from U.S. patentapplication Ser. No. 60/350,955, filed 25 Jan. 2002.

FIELD OF THE INVENTION

The invention relates to the field of treatments for insulin resistance.

BACKGROUND

Insulin resistance is a significant health challenge for a wide range ofpatients, including those with type II diabetes, metabolic obesity, andvarious liver conditions.

The picture that is emerging is one of complex multiple interactingsystems with reflex parasympathetic effects in the liver capable ofcausing more than one reaction and of triggering reactions in otherorgans.

Fasted cats develop insulin resistance immediately following acutedenervation of the liver. In such studies, the degree of reduction ofresponse to insulin was maximal after anterior plexus denervation anddid not increase further with addition of denervation of the posteriornerve plexus or bilateral vagotomy thus demonstrating that all of thenerves of relevance were in the anterior plexus.

The rapid insulin sensitivity test (RIST) was employed (Lautt et al.,Can. J. Physiol. Pharmacol. 76:1080 (1998)) is employed to avoid thecomplexity of the reaction to hypoglycemia. The RIST involves use of aeuglycemic clamp following the administration of insulin andquantitation of the response as the amount of glucose required to beinfused over the test period in order to hold arterial blood glucoselevels constant. The RIST methodology has been published in detail andhas been demonstrated in both cats and rats. It is highly reproduciblewith up to five consecutive responses being obtainable in cats and fourin rats with blood glucose levels returning to control levels betweeneach test. Insulin, glucagon, and catecholamine levels remain unchangedbetween tests.

Cats show a dose-related development of insulin resistance usingatropine (a cholinergic muscarinic receptor antagonist) that was of asimilar magnitude to that produced by surgical denervation. The dose ofatropine required to produce a full insulin resistance is 3 mg/kg (4μmol/kg) administered into the portal vein. A similar degree of insulinresistance was achieved with 10⁻⁷ mmol/kg of the M₁ muscarinic selectiveantagonist, pirenzepine, and with 10⁻⁶ μmol/kg of the M₂ selectiveantagonist, methoctramine. Although not conclusive, the data suggestthat the response may be mediated by the M₁ muscarinic receptor subtype.

Although the liver appeared to be the organ that produced the insulinresistance, it was not clear that the liver was the resistant organ. Inorder to determine the site of insulin resistance, a further series wasdone in cats that measured arterial-venous glucose responses across thehindlimbs, extrahepatic splanchnic organs, and liver. The intestine wasunresponsive to the bolus insulin administration both before and afteratropine or anterior plexus denervation or the combination of both. Thehepatic response was also not notably altered whereas the glucose uptakeacross the hindlimbs, primarily representing skeletal musde uptake, wasdecreased following atropine or hepatic parasympathetic denervation.These results indicated that interference with hepatic parasympatheticnerves led to insulin resistance in skeletal muscle.

It was further demonstrated that the same degree of resistance could beproduced by pharmacological blockade of parasympathetic nerve functionusing the muscarinic receptor antagonist, atropine. Following a meal,insulin is released from the pancreas. The presence of insulin in theblood elicits a hepatic parasympathetic reflex that results in therelease of acetylcholine in the liver that results in the generation andrelease of nitric oxide which acts to control the sensitivity ofskeletal muscle to insulin through the action of a hormone released fromthe liver, a hepatic insulin sensitizing substance (HISS) whichselectively stimulates glucose uptake and storage as glycogen in tissuesincluding skeletal muscle.

In the absence of HISS, the large muscle mass is highly resistant toinsulin and the glucose storage in skeletal muscle is severely reduced.Interruption of any part of the parasympathetic-mediated release of HISSresults in insulin resistance. This parasympathetic reflex regulation ofHISS release is an important mechanism by which the body regulatesresponsiveness to insulin and this mechanism is adjusted according tothe prandial state, that is, according to how recently there has been anoral consumption of nutrients.

In a fasted condition, HISS release in response to insulin is minimal orabsent so that if insulin is released in this situation, there is aminimal metabolic effect. Following a meal, the parasympathetic reflexmechanism is amplified so that HISS release occurs and results in themajority of the ingested glucose stored in skeletal muscle.

The consequence of lack of HISS release is the absence of HISS whichresults in severe insulin resistance, referred to as HISS-dependentinsulin resistance (“HDIR”). In this situation, the pancreas is requiredto secrete substantially larger amounts of insulin in order that theglucose in the blood is disposed of to prevent hyperglycemia fromoccurring. If this condition persists, insulin resistance will progressto a state of type 2 diabetes (non-insulin dependent diabetes mellitus)and eventually will lead to a complete exhaustion of the pancreas thusrequiring the patient to resort to injections of insulin. Thus, itappears that any condition in which the hepatic parasympathetic reflexis dysfunctional will result in insulin resistance.

It is believed that the insulin resistance that is seen in a variety ofconditions (non-insulin dependent diabetes, essential hypertension,obesity, chronic liver disease, fetal alcohol effects, old age, andchronic inflammatory diseases) represents a state of HDIRparasympathetic dysfunction. Lack of HISS would also be anticipated toresult in obesity at the early stage of the resultant metabolicdisturbance (the obese often become diabetic).

Normally after a meal the liver takes up a small proportion of glucoseand releases HISS to stimulate skeletal muscle to take up the majorityof the glucose load. In the absence of HISS, the skeletal muscle isunable to take up the majority of glucose thus leaving the liver tocompensate. The hepatic glycogen storage capacity is insufficient tohandle all of the glucose, with the excess being converted to lipidswhich are then incorporated into lipoproteins and transported to adiposetissue for storage as fat. Provision of HISS to these individuals wouldrestore the nutrition partitioning so that the nutrients are storedprimarily as glycogen in the skeletal muscle rather than as fat in theadipose tissue.

Thus, it is an object of the invention to provide a method of reducinginsulin resistance.

SUMMARY OF THE INVENTION

Nitric oxide (NO), GSH and insulin are believed to act within the liverto cause the release of a hepatic insulin sensitizing substance (HISS)into the blood. HISS controls the sensitivity of certain tissues(including skeletal muscle) to insulin. When HISS is present insignificant amounts these tissues become more sensitive to insulin andcan rapidly take up and store glucose. While the invention is notlimited to any particular mechanism, the insulin sensitizing effect ofHISS is believed to result when insulin causes HISS release and thenHISS action is imposed on skeletal muscle. The direct action of insulinon peripheral tissues is not believed to be directly altered. Similarly,the term HISS-dependant insulin resistance refers to the decreasedglucose storage effect produced by insulin as a result of lack of HISSaction, not a direct reduction in cellular response to insulin.

In an embodiment of the invention there is provided a method of reducingHISS-dependent insulin resistance in a mammalian patient sufferingtherefrom comprising administering a glutathione-increasing compound andin some instances a nitric oxide increasing compound.

Acetylcholine infused directly into the portal vein (2.5 μg/kg/min)results in a complete reversal of the insulin resistance induced bysurgical denervation. Administration of the same dose of acetylcholineintravenously produces no reversal. Intraportal administration directlytargets the liver whereas intravenous infusion bypasses the liver and isnot organ selective. This demonstration is extremely important in thatthe data suggest that the signal from the liver to skeletal muscle isblood-borne.

A small but significant reduction in insulin resistance is observedfollowing the administration of GSH alone. However, this effect isinsufficient for many therapeutic purposes and is believed to beHISS-independent. Improved insulin sensitivity is obtained when both NOand GSH are administered.

A role for nitric oxide (“NO”) in reducing insulin resistance has beenpreviously reported (Int. Pub. WO 00/19992 of Lautt). However, it hassubsequently been discovered that NO alone is insufficient to fullyovercome HISS-dependent insulin resistance in patients sufferingtherefrom and having low levels of hepatic GSH. While NO alone canincrease insulin sensitivity in such patients, the effect is notcomplete and may not be sufficient for therapeutic purposes. As usedherein, the phrase “low levels of hepatic GSH” refers to GSH levelslower than 3.5 μmol/g fresh liver when assayed after a 16 to 24 hourfast.

In an embodiment of the invention there is provided use of aglutathione-increasing compound and a nitric oxide increasing compoundin reducing HISS-dependent insulin resistance in a mammalian patientsuffering therefrom.

In an embodiment of the invention there is provided use of aglutathione-increasing compound and a nitric oxide increasing compoundin the manufacture of a medicament useful in the treatment ofHISS-dependent insulin resistance.

In an embodiment of the invention there is provided use of a compositioncomprising a glutathione-increasing compound and a nitricoxide-increasing compound in improving glucose uptake in a patientsuffering from insulin resistance.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 is a schematic illustration of the experimental protocol employedin Example 1.

FIGS. 2 a-d depict the RIST index for the control group BSO group inExample 1, as well as HISS dependent and independent components ofinsulin action in the BSO and control groups, and hepatic GSH content inthose groups.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

Surprisingly, it has been learned that the combined action ofglutathione and nitric oxide (“NO”) play an important role in theproduction of HISS in the liver. Glutathione, NO and insulin are neededfor the release of sufficient active HISS to overcome HISS-dependentinsulin resistance (“HDIR”).

HISS-dependent insulin resistance refers to the reduced uptake ofglucose by HISS-sensitive tissues in the absence of HISS or when HISSlevels are insufficient. One example of such reduced uptake is that ofskeletal muscle when observed in the presence of insulin but in theabsence of a normal hepatic parasympathetic reflex. While administrationof GSH alone or NO alone may reduce insulin resistance somewhat in sucha situation, the remaining insulin resistance (not treatable with NO orGSH alone) is HDIR and can be treated by the method disclosed herein.

While the invention is not limited to any particular mechanism, it isbelieved that the administration of a glutathione increasing compoundand an NO-increasing compound restores HISS release from the liverand/or restores HISS effect at skeletal muscle and other HISS-sensitivesites of glucose uptake.

Thus, patients suffering from insulin resistance caused by insufficientHISS activity can be treated through the administration of a hepaticglutathione-increasing compound together with a hepatic NO-increasingcompound.

It will be apparent, in light of the disclosure herein, that a number ofways of increasing hepatic glutathione and NO are specificallycontemplated and fall within the scope of the invention. For example,hepatic glutathione may be increased by increasing the rate ofglutathione synthesis in the liver, reducing the rate of glutathionedegradation (other than to form HISS) in the liver, or by providingexogenous glutathione in a form which is taken up by the liver cells. Byway of non-limiting example, the rate of glutathione synthesis in theliver may in some instances be increased using one or more compounds:(a) which stimulate enzymes involved in glutathione synthesis (but thecompounds are not reactants in the reactions producing glutathione); (b)which are reactants in the reaction producing glutathione; or (c) whichstimulate the production of one or more subsequent compounds whicheither stimulate glutathione producing enzymes or are reactants in thereaction producing glutathione. In light of the disclosure herein, oneskilled in the art could select a suitable method of increasing hepaticglutathione. Examples of glutathione-increasing compounds include:n-acetylcysteine, cysteine esters, L-2-oxothiazolidine-4-carboxolate(“OTC”), gamma glutamylcysteine and its ethyl ester, glutathione ethylester, glutathione isopropyl ester, lipoic acid, cystine, cysteine,methionine, and s-adenosylmethionine.

Similarly, hepatic NO levels can be increased by increasing the rate ofNO synthesis in the liver (such as by increasing NO synthase activity),by reducing the rate of NO degradation in the liver (other than to formHISS), or by providing exogenous NO or an exogenous carrier or precursorwhich is taken up and releases NO in the liver.

NO-increasing compounds include SIN-1 and molsidamine, and nitrosylatedforms of: N-acetylcysteine, cysteine esters,L-2-oxothiazolidine4-carboxolate (“OTC”), gamma glutamylcysteine and itsethyl ester, glutathione ethyl ester, glutathione isopropyl ester,lipoic acid, cystine, cysteine, methionine, and s-adenosylmethionine.

When nitrosylated forms of glutathione-increasing compounds areadministered, these compounds can perform the role of both a nitricoxide-increasing compound and a glutathione-increasing compound.

S-adenosylmethionine (SAMe) is a major regulator of hepatic glutathione.SAMe administration causes intrahepatic glutathione to be restored tonear-normal levels. Thus, SAMe administration (together with anNO-increasing compound) allows HISS production in patients otherwisesuffering from insufficient HISS as a result of insufficient hepaticglutathione. Thus, one method of increasing hepatic glutathione is bythe administration of SAMe together with an NO-increasing compound. Asused herein “SAMe” includes SAMe itself as well as pharmacologicallyacceptable salts thereof.

SAMe is preferably administered orally at a dose of 0.5 to 25 mg/kg, byintramuscular injection at a dose of 0.2 to 10 mg/kg, or by anotherroute of administration with an equivalent dose.

Peak glutathione levels are typically seen about 3-6 hours after oraladministration of SAMe. In one embodiment, SAMe is administered once perday. In some instances the daily oral dose is preferably taken in themorning.

The precise dose of glutathione-increasing compound and NO-increasingcompound desirable will be determined by a number of factors which willbe apparent to those skilled in the art, in light of the disclosureherein. In particular, the identity of the glutathione-increasingcompound and the NO-increasing compound and its mechanism of action (ifknown), the formulation and route of administration employed, theunstimulated level of HISS production in response to the patient'sendogenous glutathione and/or NO levels, the patient's gender, age andweight, as well as the extent of unstimulated hepatic glutathione and/orNO production and the severity of the condition to be treated should beconsidered. Where it is impractical to conduct the tests necessary todetermine the glutathione and/or NO response and/or the other factorssuch as the extent of hepatic glutathione and/or NO production, theappropriate dose can be determined through the administration of a dosesuitable for a majority of patients similar to the subject in respect ofthose factors which have been assessed, followed by monitoring of thepatient to determine if HISS production is increased. Such monitoringmay be by any suitable means including, for example, the RIST.

In one embodiment of the invention, a patient suffering from insulinresistance is treated for that condition by the administration of one ormore of S-adenosylmethionine, vitamin E, vitamin C and3-morpholinosyndnonimine and other pharmaceutically acceptableanti-oxidants, together with a NO-increasing compound.

For oral administration, in some instances the components are preferablyadministered immediately before a meal: SAMe (1 to 20 mg/kg, preferably6 mg/kg body weight), vitamin E (1 to 20 mg/kg, preferably 6 mg/kg bodyweight), vitamin C (1 to 20 mg/kg, preferably 7 mg/kg body weight), and3-morpholinosyndnonimine (“SIN-1”) (50 to 400 mg/kg, preferably 150mg/kg body weight).

In one embodiment of the invention the glutathione-increasing compoundis 8-bromo-cGMP. 8-bromo-cGMP for administration to a patient may be inany pharmaceutically acceptable carrier. In some instances intravenousadministration at a dose of 0.05 to 1 mg/kg/min infused continuouslywill be desired. In some instances a dose of 1 to 0.5 mg/kg/min will bedesired. Infusion will preferably commence near the time of a meal sothat 8-bromo-cGMP levels in the liver will peak when blood glucoselevels are elevated above pre-meal levels. In some instances8-bromo-cGMP may be continuously administered for 1 to 60 minutes. Insome instances administration for 5 to 30 minutes will be desirable.Comparable oral doses may also be employed, although in some instancesit may be desirable to administer an oral dose a greater time prior to ameal than the comparable intravenous dose would be administered.

In one embodiment, the NO-increasing compound is SIN-1. SIN-1administration to a patient may be in any suitable carrier. In someinstances, an intravenous dose of between about 1 and 25 mg/kg bodyweight administered over a 5 minute period to 1 hour will be desired. Insome instances a dose of between about 5 and 20 mg/kg body weight willbe desired. In some instances a dose of between 8 and 15 mg/kg bodyweight will be desired. In some instances the period over which the doseis administered will be adjusted to allow hepatic NO levels to rise inparallel to hepatic GSH levels.

Any suitable glutathione-increasing compound and any suitableNO-increasing compound may be employed. A glutathione-increasing orNO-increasing compound will be “suitable” if: (a) at the dose and methodof administration to the mammalian patient, it is not acutely toxic, anddoes not result in chronic toxicity disproportionate to the therapeuticbenefit derived from treatment; and (b) at the dose and method ofadministration to the mammalian patient, including the impact of asuitable dose of the other (GSH or NO) increasing compound, it reducesinsulin resistance in the patient.

In one embodiment there is provided a pharmaceutical compositionincluding a hepatic glutathione-increasing compound and a hepatic nitricoxide-increasing compound.

In one embodiment of the invention, the glutathione-increasing compoundand NO-increasing compound are preferentially targeted to the liver.Targeting to the liver can be accomplished through the use of anypharmaceutically acceptable liver targeting substance. For example, eachcompound can be bound to albumin for preferential delivery to liver;alternatively, the compounds may be incorporated into or encapsulatedwithin liposomes which are preferentially targeted to the liver.Compounds can be bound to bile salts which are selectively taken up bythe liver. In one embodiment, one or both compounds are administered ina precursor form, and the precursor is selected to be metabolised to theactive form by enzymes preferentially found in the liver.

The glutathione-increasing compound may be administered so as tomaintain a relatively constant level of the glutathione-increasingcompound in the liver at all times. In some instances it will bepreferable to administer the glutathione-increasing compound so as tohave its concentrations peak when blood glucose is high, such as after ameal, allowing HISS production and glucose uptake at that time. Wheretoxicity is a concern, it may be desirable to keepglutathione-increasing compound levels low until blood glucose levelsbecome elevated above normal levels. Alternatively, theglutathione-increasing compound may be administered so as to maintain arelatively constant level of HISS in the liver at all times.Alternatively, the glutathione-increasing compound may be administeredto have HISS concentrations peak when blood glucose is high, and toremain high for no more than about 4 to 6 hours.

In some instances, it will be desirable to administer a NO-increasingcompound and a GSH-increasing compound together with at least one otherdrug used in the treatment of diabetes, examples of which are listed inTable I. TABLE I a. Insulin and insulin analogues b. Type II Diabetesdrugs i. Sulfonylurea agents 1. First Generation   a. Tolbutamide   b.Acetohexamide   c. Tolazamide   d. Chlorpropamide 2. Second Generation  a. Glyburide   b. Glipizide   c. Glimepiride ii. Biguanide agents 1.metformin iii. Alpha-glucosidase inhibitors 1. Acarbose 2. Miglitol iv.Thiazolidinedione Agents (insulin sensitizers) 1. Rosiglitazone 2.Pioglitazone 3. Troglitazone v. Meglitinide Agents 1. Repaglinide c.Phosphodiesterase Inhibitors i. Anagrelide ii. Tadalafil iii.Dipyridamole iv. Dyphylline v. Vardenafil vi. Cilostazol vii. Milrinoneviii. Theophylline ix. Sildenafil x. Caffeine d. CholinesteraseInhibitors i. Donepezil ii. Tacrine iii. Edrophonium iv. Demecarium v.Pyridostigmine vi. Phospholine vii. Metrifonate viii. Neostigmine ix.Galanthamine x. Zanapezil e. Cholinergic Agonists i. Acetylcholine ii.Methacholine iii. Bethanechol iv. Carbachol v. Pilocarpine hydrochloride

In one embodiment of the invention there is provided a method ofreducing insulin resistance in a mammalian patient having lower thannormal hepatic glutathione levels. In this embodiment the methodcomprises: selecting a patient suffering from insulin resistance,determining if hepatic glutathione levels are lower than normal in thepatient, and administering a compound which increases hepaticglutathione.

As used herein, the phrase “lower than normal hepatic glutathionelevels” means hepatic glutathione levels lower than those observed in anaverage healthy individual of the same gender, age, weight, fed-state,and blood sugar level.

The patient is preferably a mammal. In one embodiment the patient is ahuman. In another embodiment the patient is a domestic animal such as acat, dog or horse.

EXAMPLES

Summary: GSH depletion was induced in 5 week old Wistar rats usingbuthionine sulfoximine (BS), 2 mmol/kg bw, i.p. for 20 days. Controlrats were injected with an equal volume of saline. Insulin sensitivitywas measured using a euglycemic clamp, the Rapid Insulin SensitivityTest—RIST. In both the saline group (SG) and BSO group (BG) a controlRIST followed by a RIST post-L-NAME (1 mg/kg i.p.v.), a NOS competitiveantagonist, and a RIST post-SIN-1 (5 mg/kg i.p.v.), a NO donor, wereperformed. The liver was removed for further determined of hepatic GSHcontent using the GSH peroxidase-reductase assay. BG showed a reductionof 48.3±6.9% in hepatic GSH content compared to SG. The control RIST,248.0±15.8 mg glucose/kg for SG, was impaired in the BG (158.4±12.2 mgglucose/kg; p<0.01). After L-NAME the SG RIST was 129.0±1.9 mgglucose/kg and the BG RIST was 109.0±9.1 mg glucose/kg. SIN-1administration only restored insulin action in the SG (246.0±11.5 mgglucose/kg). HISS action, quantified by subtracting the post L-NAME RISTfrom the control RIST, was 114.0±15.8 mg glucose/kg for SG and only49.3±8.6 mg glucose/kg for BG (p<0.01). HISS dependent insulinresistance appears to be related to impaired levels of GSH in the liver,leading to a compromised HISS secretion/release.

SIN-1 is a NO donor. L-NAME and L-NMMA are nitric oxide synthase (NOS)antagonists.

SIN-1 reverses HISS inhibition after NOS blockade.

SIN-1 produces NO and O₂ ²⁻ simultaneously. Nitrosatin of GSH by NO/O₂²⁻ to produce GSNO is efficient. GSNO is apparently essential for HISSrelease. Nitroprusside is not able to restore HISS action after NOSblockade.

Example 1

The overall experimental protocol is shown in FIG. 1. Male Wistar rats(8 weeks) were anaesthetized with sodium pentobaribtial (65 mg/kg). Bodytemperature was maintained at 37±0.5□C. An arterial-venous loop (carotidartery and internal jugular vein) was employed. A catheter was insertedinto the portal vein.

RIST (Rapid Insulin Sensitivity Test): Blood samples (25 μl) forarterial glucose analysis were taken from the arterial-venous loop. Astable glucose baseline was established. Insulin (50 mU/kg i.v.) wasinfused over 5 minutes. Arterial blood samples were taken every 2minutes. Glucose was adjusted to maintain euglycemia. RIST index=mgglucose/kg infused during RIST.

CONTROL GROUP: Control group (n=6): L-NAME (1 mg/kg i.p.v.) reduces theRIST index from 260.2±15.6 mg glucose/kg to 121.2±12.8 mg glucose/kg(52.3±5.8% inhibition). SIN-1 (5 mg/kg i.p.v.) restores insulin responsewith a RIST index of 258.1±18.5 mg glucose/kg. ***=p<0.001.

BSO GROUP: BSO group (n=5): The control RIST index was 158.4±12.2 mgglucose/kg. Intraportal administration of L-NAME (1 mg/kg) reducedsignificantly the RIST index to 109.8±9.1 mg glucose/kg i.p.v.Administration of SIN-1 did not reverse the RIST index to controlvalues. *=p<0.05; ns=non significant.

INSULIN ACTION: HISS-dependent and HISS-independent components ofinsulin action in BSO and control groups. HISS-independent componentsare not different in both groups. The direct glucose disposal action ofinsulin is unaltered by GSH depletion. HISS is significantly reduced inBSO group (49.3±8.56 mg glucose/kg) compared to control group(138.9±22.8 mg glucose/kg) corresponding to a decrease of 64.4% of HISSaction. **=p<0.01; ns=non significant.

HEPATIC GSH CONTENT: Hepatic glutathione content in BSO (n=5) andcontrol (n=6) groups. In control group hepatic GSH content wassignificantly higher (5.66±0.1 μmol/g fresh liver) than in BSO group(2.96±0.4 μmol/g fresh liver). Hepatic GSH content was decreased in48.3±6.9% in BSO group. ***=p<0.001.

The results are shown in FIG. 2(a-d).

An impaired content of hepatic GSH appears to lead to insulinresistance. The HISS-independent component of insulin action is notaltered in the GSH depleted rats. The HISS dependent component ofinsulin action is decreased significantly in BSO group compared withcontrol group. HISS synthesis/release is dependent on the presence ofGSH in the liver.

Example 2

A model of low hepatic GSH and NO production is produced by overnightfasting. Normal hepatic levels of GSH are reduced by an overnight fast.SIN-1 at doses from 2.5 to 10 mg/kg i.p.v. do not reverse fastinginduced HDIR but prior administration of glutathione esters raiseshepatic GSH levels to those seen in fed animals and restores the abilityof the NO donor SIN-1 to serve as the permissive signal that allowsinsulin to cause HISS release thereby reversing fasting-induced HDIR.GSH administration by itself results in minor reversal of HDIR whichonly becomes therapeutically significant after the addition of the NOdonor.

Thus, there has been provided a method of reducing insulin resistance.

Publications relating to the material described herein include:

Dowell, F. J et al.: Eur. J. Pharmacol. 379(23):175-182 (1999); Young,M. E., Leighton, B.: Biochem. J. 329:73 (1998); Xie, H., Lautt, W. W.:Am. J. Physiol. 270:E858 (1996); Lautt, W. W. et al.: Can. J. Physiol.Pharmacol. 76:1 (1998); Sadri, P., Lauft, W. W.: Am. J. Physiol.277:G101 (1999); Modan, M., Halkin, H., Almog, S.: Journal of ClinicalInvestigation. 75:809 (1985); DeMattia, G. et al.: Metabolism 47:993(1998); Czech, M. P. et al.: Metabolism. 27:1987 (1978); Xie, H., Lautt,W. W.: Am. J. Physiol. 270:E858 (1996); Sadri, P., Lauft, W. W.: Am. J.Physiol. 277:G1 (1999); Khamaisi, M., Kavel, O., Rosenstock, M., Porat,M., Yull, M., Kaiser, N., Rudich A.: Biochem. J. 349:579 (2000); Lauft,W. W., Wang, X., Sadri, P., Legare, D., Macedo, M. P.: Can. J. Physiol.Pharmacol. 76:1 (1998); Marinho, H. S., Baptista, M., Pinto, R. E.:Biochem. Biophys. Acta. 1360:157 (1997); Schrammel, A., Pfeiffer, S.,Schmidt, K., Koesling, D., Mayer, B.: Mol. Pharmacol. 54:207 (1998).

1. Use of a glutathione-increasing compound and a nitric oxideincreasing compound in the manufacture of a medicament useful in thetreatment of insulin resistance.
 2. Use of a glutathione-increasingcompound and a nitric oxide-increasing compound in improving glucoseuptake in a patient suffering from insulin resistance.
 3. Use of claim 1wherein the insulin resistance is hepatic insulin sensitizing substance(“HISS”)-dependent insulin resistance.
 4. Use of claim 1 wherein theglutathione-increasing compound is at least one of N-acetylcysteine,cysteine esters, L-2-oxothiazolidine-4-carboxolate (“OTC”), gammaglutamylcysteine and its ethyl ester, glutathione ethyl ester,glutathione isopropyl ester, lipoic acid, cystine, cysteine, methionine,and S-adenosylmethionine (“SAMe”).
 5. Use of claim 1 wherein the nitricoxide-increasing compound is at least one of SIN-1, molsidamine,nitrosylated N-acetylcysteine, nitrosylated cysteine esters,nitrosylated L-2-oxothiazolidine-4-carboxolate (NOTC), nitrosylatedgamma glutamylcystein and its ethyl ester, nitrosylated glutathioneethyl ester, nitrosylated glutathione isopropyl ester, nitrosylatedlipoic acid, nitrosylated cysteine, nitrosylated cystine, nitrosylatedmethionine, or nitrosylated S-adenosylmethionine.
 6. A pharmaceuticalcomposition comprising a hepatic glutathione increasing compound and ahepatic nitric oxide-increasing compound.
 7. A pharmaceuticalcomposition comprising at least one of nitrosylated N-acetylcysteine,nitrosylated cysteine esters, nitrosylatedL-2-oxothiazolidine-4-carboxolate (NOTC), nitrosylated gammaglutamylcystein and its ethyl ester, nitrosylated glutathione ethylester, nitrosylated glutathione isopropyl ester, nitrosylated lipoicacid, nitrosylated cysteine, nitrosylated cystine, nitrosylatedmethionine, or nitrosylated S-adenosylmethionine.
 8. The pharmaceuticalcomposition of claim 6 further including a pharmaceutically acceptableantioxidant.
 9. A method of reducing insulin resistance in a mammalianpatient having lower than normal hepatic glutathione levels, said methodcomprising: selecting a patient suffering from insulin resistance;determining if hepatic glutathione levels are lower than normal in thepatient; and administering a compound which increases hepaticglutathione and a compound which increases hepatic nitric oxide.
 10. Amethod of reducing insulin resistance in a mammalian patient comprisingadministering a compound which increases hepatic glutathione and acompound which increases hepatic nitric oxide (“NO”).
 11. Thecomposition of claim 6 further including a pharmaceutically acceptableliver-targeting substance.
 12. The method of claim 9 wherein the insulinresistance is HISS-dependent insulin resistance (“HDIR”).
 13. The methodof claim 12 wherein the hepatic glutathione-increasing compoundadministered causes an increase in hepatic glutathione synthesis. 14.The method of claim 10 wherein the glutathione-increasing compound is atleast one of N-acetylcysteine, cysteine esters,L-2-oxothiazolidine-4-carboxolate (“OTC”), gamma glutamylcysteine andits ethyl ester, glutathione ethyl ester, glutathione isopropyl ester,lipoic acid, cystine, cysteine, methionine, and S-adenosylmethionine(“SAMe”).
 15. The method of claim 10 wherein the nitric oxide-increasingcompound is at least one of SIN-1, molsidamine, nitrosylatedN-acetylcysteine, nitrosylated cysteine esters, nitrosylatedL-2-oxothiazolidine-4-carboxolate (NOTC), nitrosylated gammaglutamylcystein and its ethyl ester, nitrosylated glutathione ethylester, nitrosylated glutathione isopropyl ester, nitrosylated lipoicacid, nitrosylated cysteine, nitrosylated cystine, nitrosylatedmethionine, or nitrosylated S-adenosylmethionine.
 16. The method ofclaim 1 wherein the glutathione-increasing compound is administeredorally.
 17. The method of claim 1 wherein the glutathione-increasingcompound is administered by intravenous injection.
 18. The method ofclaim 1 wherein the glutathione-increasing compound is 8-bromo-cGMP. 19.The method of claim 1 wherein the compound which increases hepatic NO isadministered orally.
 20. The method of claim 1 wherein the compoundwhich increases hepatic NO is administered by intravenous injection. 21.The method of claim 1 wherein the compound which increases nitric oxideis SIN-1.
 22. The method of claim 1 wherein the compound which increaseshepatic NO is molsidamine.
 23. The method of claim 1 further includingadministering a pharmaceutically acceptable anti-oxidant.
 24. The methodof claim 1 wherein the patient suffers from at least one of non-insulindependent diabetes, essential hypertension, metabolic obesity, chronicliver disease, fetal alcohol effects, old age and a chronic inflammatorydisease.
 25. The method of claim 1 wherein the patient is a human. 26.Use of claim 2 wherein the insulin resistance is hepatic insulinsensitizing substance (“HISS”)-dependent insulin resistance.
 27. Use ofclaim 2 wherein the glutathione-increasing compound is at least one ofN-acetylcysteine, cysteine esters, L-2-oxothiazolidine-4-carboxolate(“OTC”), gamma glutamylcysteine and its ethyl ester, glutathione ethylester, glutathione isopropyl ester, lipoic acid, cystine, cysteine,methionine, and S-adenosylmethionine (“SAMe”).
 28. Use of claim 2wherein the nitric oxide-increasing compound is at least one of SIN-1,molsidamine, nitrosylated N-acetylcysteine, nitrosylated cysteineesters, nitrosylated L-2-oxothiazolidine-4-carboxolate (NOTC),nitrosylated gamma glutamylcystein and its ethyl ester, nitrosylatedglutathione ethyl ester, nitrosylated glutathione isopropyl ester,nitrosylated lipoic acid, nitrosylated cysteine, nitrosylated cystine,nitrosylated methionine, or nitrosylated S-adenosylmethionine.
 29. Thepharmaceutical composition of claim 7 further including apharmaceutically acceptable antioxidant.
 30. The composition of claim 7further including a pharmaceutically acceptable liver-targetingsubstance.